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Streptococcus mutans  is an important pathogen in the human oral biofilm. It expresses virulent behaviors that are linked to its genetic competence regulon, which is controlled by  comX . Expression of  comX  is modulated by two diffusible signaling peptides, denoted CSP and XIP, and by other environmental cues such as pH and oxidative stress. The sensitivity of  S. mutans  competence to environmental inputs that may vary on microscopic length scales raises the question of whether the biofilm environment creates microniches where competence and related phenotypes are concentrated, leading to spatial clustering of  S. mutans  virulence behaviors. We have used two-photon microscopy to characterize the spatial distribution of  comX  expression among individual  S. mutans  cells in biofilms. By analyzing correlations in  comX  activity, we test for spatial clustering that may suggest localized competence microenvironments. Our data indicate that both competence-signaling peptides diffuse efficiently through the biofilm. XIP elicits a population-wide response. CSP triggers a Poisson-like, spatially random  comX  response from a subpopulation of cells that is homogeneously dispersed. Our data indicate that competence microenvironments if they exist are small enough that the phenotypes of individual cells are not clustered or correlated to any greater extent than occurs in planktonic cultures.

Spatial Correlations and Distribution of Competence Gene Expression in Biofilms of Streptococcus mutans